Photodynamic therapy of extracellular matrix stimulates endothelial cell growth by inactivation of matrix-associated transforming growth factor-beta

Photodynamic therapy (PDT), the production of cytotoxic free-radical moieties by light activation of photosensitizer dyes, is a novel approach to inhibit experimental intimal hyperplasia. Local eradication of vascular cells with this method in vivo is followed by expedient reendothelialization, and PDT of extracellular matrix (ECM) in vitro stimulates endothelial cell (EC) growth. This in vitro study explored one possible mechanism underlying these findings by investigating the effects of PDT on matrix-associated transforming growth factor-beta (TGF-beta), a potent inhibitor of EC growth. The ECM deposited by EC on tissue culture plates contained 85.4 +/- 10.2 pg/10 cm(2) of TGF-beta, as measured by an ELISA. In contrast, after PDT of ECM, levels of TGF-beta could be barely be detected (0.2 +/- 0.5 pg/10 cm(2)). The functional consequence of this observation was demonstrated by the finding that PDT of plates coated with a fibronectin-TGF-beta complex stimulated EC mitogenesis (102.3% +/- 19.3%, p < 0.0005) compared with the untreated control (44.1% +/- 13.5%). The inhibitory effect of ECM-associated TGF-beta on EC was further delineated by blocking its activity with a specific antibody. Whereas the antibody did not affect EC mitogenesis on PDT-treated matrix or matrix-free plates (101% +/- 8.8%, 105.6% +/- 9.8%), EC mitogenesis growing on ECM was significantly enhanced (125.9% +/- 17.5%, p < 0.05). Finally, SDS-PAGE analysis of PDT-treated TGF-beta in solution demonstrated that the PDT-mediated loss of TGF-beta activity was not associated with changes in its molecular weight. These data demonstrate that increased EC proliferation on PDT-treated matrix is, at least in part, mediated by inactivation of TGF-beta. PDT-removal of this EC growth inhibitor in the intima provides a mechanism by which PDT of the vascular wall could potentiate endothelial regrowth, a factor which may promote proper healing and result in the inhibition of intimal hyperplasia.